One of major objects of researches in organic chemistry is to create highly efficient chemical catalysts. In development of catalysts, besides determination of whether a test substance functions as a highly active catalyst, it is essential to optimize reaction conditions. Conventionally for developing catalysts, a means has been used in which advance of a catalytic reaction is monitored over period of time by high performance liquid chromatography or the like. However, a huge number of chemical substances are created due to recent progress of combinatorial techniques, and as a result, it has become difficult to conduct a screening for chemical substances having applicability as catalysts by applying the conventional means. Moreover, even if chemical substances having applicability as catalysts are chosen, it is still impossible to efficiently perform development of catalysts including optimization of reaction conditions by using the conventional means. From such viewpoints, it has been desired to provide a means that enables efficient screening for chemical substances having applicability as catalysts and convenient selection of optimum reaction conditions in a short time.
The reaction of Michael addition is widely used in the field of organic synthesis as a fundamental carbon-carbon bond formation reaction, and it is desired to develop a system capable of efficiently advancing the reaction of Michael addition. If efficient screening for catalytic substances used in the reaction of Michael addition and efficient determination of conditions for the reaction system are achieved, it is expected that the development of the reaction system of Michael addition, which is currently carried out for every test substance on trial and error basis, can be remarkably progressed.
Direct fluorescence visualization of intracellular localizations and dynamic behaviors of proteins in living cells and tissues is extremely important for elucidation of physiological functions of the proteins, and techniques using a protein fused with GFP (Green Fluorescent Protein) are widely used in recent years. However, behavior of a target protein may possibly not be correctly monitored with GFP due to a problem concerning a molecular size or the like of GFP itself. Accordingly, a means has been desired for achieving specific and highly sensitive fluorescence visualization of a target protein based on introduction of a fluorescence tag having a smaller molecular size. The inventors of the present invention found that a compound having 7-hydroxycoumarin as a fluorophore and two maleimide groups reacted with a peptide having two adjacent cysteine residues in the same molecule, and revealed that the compound was substantially non-fluorescent before the reaction, whilst it came to emit intense fluorescence after the reaction, and thereby the compound was useful as a technique for introducing a fluorescence tag into a peptide (Abstract of Photochemistry Symposium, Sep. 12 to 14, 2005). However, excitation wavelength of the aforementioned compound having 7-hydroxycoumarin and two maleimide groups is in the ultraviolet region, and accordingly the compound may sometimes causes a problem of cytotoxicity. Moreover, from a viewpoint of application to a biological system, improvement of reaction selectivity to a peptide having two adjacent cysteine residues in a molecule was another object. Therefore, it has been desired to develop a technique for highly selective introduction of a fluorescence tag into a protein, and for excitation with a visible light that is free from cytotoxicity with excitation.